Certain heterocyclic thiomethyl derivatives of 7-(dihalogenated phenylthioacetamido)cephalosporanic acids useful in the treatment of infections caused by _Streptococcus faecalis

ABSTRACT

7-(2,5-Dichloro or 3,4-dichlorophenyl) cephalosporanic acids or derivatives thereof in which the acetoxy group is replaced by hydroxyl or certain heterocyclic thio groups are useful in the treatment of infections caused by Streptococcus faecalis.

This is a division of application Ser. No. 35,035, filed May 1, 1979,now U.S. Pat. No. 4,261,991.

This invention relates to a method of treating a bacterial infectioncaused by strains of Streptococcus faecalis using certain7-(dihalogenated phenylthioacetamido) cephalosporanic acids andderivatives thereof. Also contemplated are the novel compounds useful insaid method.

In its process aspect, the invention sought to be patented comprehends amethod or treating a bacterial infection caused by a strain ofStreptococcus faecalis which comprises administering to a warm-bloodedhost afflicted with said infection an effective amount of a compound ofthe formula: ##STR1## wherein R is 2,5-dichlorophenyl or3,4-dichlorophenyl; and Y is hydroxy, acetoxy, or a heterocyclic thiomoiety --S--X wherein X is a heterocyclic group of the formula ##STR2##or a salt thereof with a non-toxic, pharmaceutically acceptable cation,provided that when R is 3,4-dichlorophenyl, Y cannot be hydroxy oracetoxy.

In subgeneric aspects, the invention includes the following embodiments:

(i) compounds of Formula I wherein R is 2,5-dichlorophenyl;

(ii) compounds of Formula I wherein R is 2,5-dichlorophenyl and Y isacetoxy;

(iii) compounds of Formula I wherein R is 2,5-dichlorophenyl and Y is aheterocyclic thio moiety as defined supra; and

(iv) compounds of Formula I wherein R is 3,4-dichlorophenyl and Y is aheterocyclic thio moiety as defined supra.

The compounds of Formula I are either known or are prepared byconventional methods from known intermediates. U.S. Pat. No. 3,335,135describes the compounds of Formula I wherein R is 2,5-dichlorophenyl or3,4-dichlorophenyl and Y is acetoxy. U.S. Pat. No. 4,056,676 describesthe compound of Formula I wherein R is 2,5-dichlorophenyl or3,4-dichlorophenyl and Y is acetoxy, 5-methyl-1,3,4-thiadiazol-2-ylthio,or 1-methyl-1H-tetrazol-5-ylthio.

The compounds of Formula I wherein Y is a heterocyclic thio group ingeneral are prepared from a 7-(2,5- or3,4-dichlorophenylacetamido)cephalosporanic acid by nucleophilicdisplacement of the acetoxy group by an appropriate heterocyclic thiogroup. This reaction can be performed in known manner, either in anaqueous medium by reacting a salt (e.g., sodium) of the desiredcephalosporanic acid derivative with the sulfur nucleophile at pH 5-8 atan elevated temperature (35°-70° C.) (see, for example, U.S. Pat. No.3,278,531 and J. Cocker et al., J. Chem. Soc., 1965, 5015), or in anorganic solvent under anhydrous conditions by reacting thecephalosporanic acid derivative with the sulfur nucleophile at anelevated temperature (50°-140° C.). (see, for example, U.S. Pat. No.4,144,391.) Other methods of preparation can be employed, such as byN-acylating the appropriate7-amino-3-[(heterocyclic)-methylthio]-3-cephem-4-carboxylic acid usingmethods conventional in the art of cephalosporin chemistry.

The compounds of Formula I may be utilized in their acid form or in theform of a salt formed by the reaction of the free carboxyl group with asuitable base. The base must have sufficient strength to neutralize theacid, and the cation derived from the base must be non-toxic andacceptable for pharmaceutical purposes. Suitable cations will beapparent to those skilled in the art. Examples are sodium, lithium,potassium, ammonium, and substituted ammonium (e.g., methyl ammonium orethyl ammonium). Less soluble salts, e.g., calcium, barium, procaine,quinine, cyclohexylbis (methyl amine) or dibenzyl ethylene diamine salt,can also be employed.

In its composition of matter aspect, the invention contemplates thenovel compounds of Formula I wherein R is 2,5-dichlorophenyl or3,4-dichlorophenyl and Y is a heterocyclic thio moiety of the formula--S--X where X is a heterocyclic group of the formula: ##STR3## or asalt thereof with a non-toxic, pharmaceutically acceptable cation. Insubgeneric aspects, the invention includes the following embodiments:

(i) the compound of Formula I wherein R is 2,5-dichlorophenyl; and

(ii) the compound of Formula I wherein R is 3,4-dichlorophenyl.

The compounds of Formula I are highly effective in inhibiting the growthof strains of Streptomyces faecalis, as evidenced by standard agardilution susceptability tests. Streptomyces faecalis is classified as aspecies of Group D β-hemolytic streptococci, also known as enterococci.Infections caused by Group D β-hemolytic streptococci, in particularStreptomyces faecalis, are generally known to be resistant tocephalosporin antibiotics. The infections caused by Group D streptococcicommonly involve the endocardium (endocarditis), the urinary tract, theintestines, the biliary tract, postoperative wounds, and interabdominalabscesses. Endocarditis is clinically a very important disease and isfrequently difficult to treat by antibiotic therapy. The disease iscommonly treated by infusion of penicillin G in combination withstreptomycin or vancomycin.

For the treatment of Streptomyces faecalis infections, a compound ofFormula I is administered parenterally, preferably by intramuscularinjection (IM) or intravenous injection (IV). The dose to beadministered will depend on various factors including the size and ageof the host, the nature and severity of the disease being treated, andthe particular physical well-being of the individual patient. Generally,for a 70 kg host, the compound is administered in an amount from about 1to about 12 g. per day (IV or IM). The effective amount of the compoundcan be administered in divided doses, e.g., 0.25 to 6 g. given two tofour times each day.

Injectable formulations containing a compound of Formula I can beprepared in the same manner as with other injectable cephalosporins, andsuitable aqueous formulations containing pharmaceutically acceptableexcipients will be known to those skilled in the art.

The compounds of Formula I may be used in combination with other knownantibiotic agents, such as streptomycin and vancomycin, and such usewill be apparent to those skilled in the art.

The following examples are illustrative of the manner of making andusing the tangible embodiments of this invention:

EXAMPLE 1

A mixture of 2,5-dichlorophenylthioacetic acid (2.37 g, 10 mM) andoxalyl chloride (5ml.) in benzene (150 ml.), with one drop ofdimethylformamide, is heated at 100° C. (steam bath) for one-half hour.Solvent is removed in vacuo leaving a residual oil. The oil, dissolvedin acetone (50 ml.) is added dropwise at 0° to a solution of deacetyl7-aminocephalosporanic acid (2.57 g., 11 mM) and sodium bicarbonate(NaHCO₃) (2.57 g.) in water (100 ml.) and acetone (50 ml.). The mixtureis stirred for one hour, after which acetone is removed in vacuo (cold).The aqueous residue is added to ethyl acetate (100 ml.) at -10° C., andthe mixture is stirred vigorously. The pH is adjusted to 2.5 withconcentrated HCl. The mixture is quickly filtered. The filtrate is thenseparated into two phases. The aqueous phase is extracted withethylacetate (50 ml.) and the organic phase is combined with theprevious organic phase. The combined organic phases are dried overanhydrous Na₂ SO₄, charcoal is added, and the mixture is allowed toremain one hour in the cold. The mixture is filtered through a talc padand the filtrate is evaporated in vacuo.7-[2-[(2,5-Dichlorophenyl)thio]acetamido]-3-hydroxymethyl-3-cephem-4-carboxylicacid is obtained as white crystals from the reduced solvent.

IR (Nujol moll): 1760 cm⁻¹ (β-lactam carbonyl); NMR (DMSOd₆): δ3.6 (2H,S, 2CH₂), 3.95 (2H, S, side chain CH₂), 4.35 (2H, S, 3--CH₂ CH₂), 5.1(1H, d, H₆), 5.7 (1H, d of d, H₇), 7.1-7.6 (3H, m, aromatic H), 9.3 (1H,d, amide NH); Titration: Mol. wt. 446.5 (Theory 449.3); λ_(max) =254(ε=16,000)

EXAMPLE 2

A mixture of3-(acetoxymethyl)-7-[2-[(2,5-dichlorophenyl)thio]acetamido]-3-cephem-4-carboxylicacid, sodium salt (255 mg, 0.5 mM], 3-mercapto-1,2,5-thiadiazole, sodiumsalt (1 mM) and pH 6.4 phosphate buffer (20 cc) is heated for 8 hours at65° C. Upon cooling, there is precipitated a cream-colored solid whichis collected by filtration and dried. Yield of7-[2-[(2,5-dichlorophenyl)-thio]acetamido]-3-[(1,2,5-thiadiazolyl)thiomethyl]-3-cephem-4-carboxylicacid, as the sodium salt: 59 mg.

    ______________________________________                                        Analysis     Theory        Found                                              ______________________________________                                        C            37.83%        33.16%                                             H            2.29          2.46                                               N            9.80          9.63                                               ______________________________________                                    

UV Analysis

λ_(max) =256 (ε=17,600).

EXAMPLE 3

A mixture of3-(acetoxymethyl)-7-[2-[(2,5-dichlorophenyl)thio]acetamido]-3-cephem-4-carboxylicacid (667 mg, 1.3 mM), 5-amino-1,3,4-thiadiazole-2-thiol (173 mg, 1.3mM), sodium bicarbonate (109 mg, 1.3 mM), and pH 6.4 buffer (25 cc.) isheated at 65° C. for 8 hours. Upon cooling there is precipitated a solidwhich is collected by filtration and dried. Yield of3-[(5-amino-1,3,4-thiadiazol-2-yl)thiomethyl]-7-[2-[(2,5-dichlorophenyl)thio]acetamido]-3-cephem-4-carboxylicacid, as the sodium salt: 240 mg.

    ______________________________________                                        Analysis     Theory        Found                                              ______________________________________                                        C            38.30%        38.5%                                              H            2.68          2.97                                               N            12.41         12.58                                              ______________________________________                                    

UV Analysis

λ_(max) =255 (ε=17,700).

EXAMPLE 4

A mixture of3-acetoxymethyl-7-[2-[(2,5-dichlorophenyl)thio]acetamido]-3-cephem-4-carboxylicacid (513 mg, 1 mM), 2-mercapto-1,3-oxazole (126 mg, 1.25 mM), sodiumbicarbonate (105 mg, 1.25 mM), and pH 6.4 buffer (25 cc.) is heated at67° C. for 8 hours. After cooling, the mixture is extracted with ethylacetate. The ethyl acetate extract is dried over magnesium sulfate andevaporated to dryness to afford a residue (577 mg) which is trituratedwith warm ethyl acetate to give7-[2-[(2,5-dichlorophenyl)thio]acetamido]-3-[(2-oxazolyl)thiomethyl]-3-cephem-4-carboxylicacid, as the sodium salt. Yield: 118 mg of dried product.

    ______________________________________                                        Analysis     Theory        Found                                              ______________________________________                                        C            42.86%        42.99%                                             H            2.84          3.04                                               N            7.89          7.62                                               ______________________________________                                    

UV Analysis

λ_(max) =255 (ε=19,200).

EXAMPLE 5

A mixture of3-acetoxymethyl-7-[2-[(2,5-dichlorophenyl)thio]acetamido]-3-cephem-4-carboxylicacid, sodium salt (513 mg. 1 mM),3-mercapto-4,5-dihydro-6-hydroxy-4-methyl-5-oxo-as-triazine hydrate (221mg., 1.25 mM), sodium carbonate (105 mg, 1.25 mM), and pH 6.4 buffer (25cc.) is warmed with slow stirring. A homogeneous reaction mixture formswithin one hour.

The mixture is heated at 65° C. for 18 hours. Upon cooling there isformed a white solid which is collected by filtration and dried. Yieldof7-[2-[(2,5-dichlorophenyl)thio]acetamido]-3-[(4,5-dihydro-6-hydroxy-4-methyl-5-oxo-as-triazin-3-yl)thiomethyl]-3-cephem-4-carboxylicacid, as the sodium salt: 240 mg.

    ______________________________________                                        Analysis     Theory        Found                                              ______________________________________                                        C            39.22%        40.19%                                             H            2.63          3.01                                               N            11.43         8.99                                               ______________________________________                                    

UV Analysis

λ_(max) =254 (ε=21,600).

EXAMPLE 6

A mixture of3-acetoxy-7-[2-[(2,5-dichlorophenyl)thio]acetamido]-3-cephem-4-carboxylicacid, sodium salt (465 mg, 0.91 mM),3-methyl-5-mercapto-1,2,4-thiadiazole (165 mg, 1.25 mM), sodiumbicarbonate (105 mg, 1.25 mM), and pH 6.4 buffer (25 cc.) is heated at65°-70° C. for 18 hours. Upon cooling, there is precipitated a solidwhich is collected by filtration and dried. Yield of7-[2-[(2,5-dichlorophenyl)thio]acetamido]-3-[(3-methyl-1,2,4-thiadiazolyl)thiomethyl]-3-cephem-4-carboxylicacid, as the sodium salt: 180 mg.

    ______________________________________                                        Analysis     Theory        Found                                              ______________________________________                                        C            38.98%        40.95%                                             H            2.58          3.23                                               N            9.57          7.01                                               ______________________________________                                    

EXAMPLE 7

A mixture of3-acetoxymethyl-7[2[(2,5-dichlorophenyl)thio]acetamido]-3-cephem-4-carboxylicacid (1.54 g, 3 mM), 2-methyl-5-mercapto-1,3,4-oxadiazole (417 mg, 3.6mM), sodium bicarbonate (303 mg., 3.6 mM) in water (20 ml.) issonnicated and then heated for 30 minutes at 64° C. to afford a clearsolution. The solution is cooled to room temperature, diluted with water(5 ml.), adjusted to pH 6.5 (from pH 6.0), and allowed to stir overnightat 63° C. Analysis of the reaction mixture by tlc on alumina gel using10:3 ether--(3:1acetic acid-water) after 18 hours showed streaking andone major spot. After 20 hours, the solution is cooled to roomtemperature. Ethyl acetate (40 ml.) is added, and the resulting mixtureis cooled to 0° C. and adjusted to pH 2.4 (from pH 7.4) with 5%hydrochloric acid. The layers are separated. The aqueous layer isextracted once with ethylacetate. The ethyl acetate phases are combined,washed with water and saturated sodium chloride solution, and dried overanhydrous sodium sulfate. The sodium sulfate is removed by filtrationand the solvent is removed by evaporation. Yield of yellow foam: 1.64 g.

The crude product is purified as follows: To a 300-mg. sample (0.547mM), dissolved in 8 ml. of methanol, is added dropwise 0.55 ml. of 1 Mlithium acetate. The solution is "scratched" and placed in arefrigerator overnight. The precipitate which forms is collected, washedwith ether, and dried. Yield of7-[2-[(2,5-dichlorophenyl)thio]acetamido]-3-[(5-methyl-1,3,4-oxadiazol-2-yl)thiomethyl]-3-cephem-4-carboxylic acid,as the lithium salt: 172 mg.

    ______________________________________                                        Analysis     Theory        Found                                              ______________________________________                                        C            41.23%        40.22%                                             H            2.73          3.26                                               N            10.12         7.01                                               Cl           12.81         12.11                                              S            17.38         12.32                                              ______________________________________                                    

UV Analysis

λ_(max) =253 (ε=14,278)

EXAMPLE 8

The in vitro activities of representative compounds of Formula I areassessed and demonstrated by the agar dilution method. The results areshown in Table 1 where the activity is measured by the MIC values,defined as the concentration of the compound capable of inhibiting thegrowth of the test organism.

                  TABLE 1                                                         ______________________________________                                        In Vitro Antibacterial Activity Against Eleven                                Strains of Streptococcus faecalis                                              ##STR4##                                                                     R           Y               MIC μg/ml*                                     ______________________________________                                        2,5-diCl phenyl                                                                            ##STR5##       1.29                                              2,5-diCl phenyl                                                                            ##STR6##       1.37                                              2,5-diCl phenyl                                                                            ##STR7##       1.88                                              2,5-diCl phenyl                                                                            ##STR8##       1.88                                              2,5-diCl phenyl                                                                            ##STR9##       2.00                                              2,5-diCl phenyl                                                                            ##STR10##      2.13                                              2,5-diCl phenyl                                                                            ##STR11##      3.11                                              2,5-diCl phenyl                                                                           OH              3.11                                              2,5-diCl phenyl                                                                            ##STR12##      3.76                                              2,5-diCl phenyl                                                                            ##STR13##      4.26                                              3,4-diCl phenyl                                                                            ##STR14##      1.76                                              3,4-diCl phenyl                                                                            ##STR15##      4.83                                              ______________________________________                                         *Geometric mean MIC as determined by the tube dilution method against         eleven strains of Streptococcus faecalis.                                

What is claimed is:
 1. A compound of the formula ##STR16## wherein R is2,5-dichlorophenyl or 3,4-dichlorophenyl, and Y is a heterocyclic thiomoiety of the formula --S--X wherein X is a heterocyclic group selectedfrom ##STR17## or a salt thereof with a non-toxic, pharmaceuticallyacceptable cation.
 2. A compound as defined in claim 1 wherein R is2,5-dichlorophenyl.
 3. The compound defined in claim 2 wherein thecompound is7-[2[(2,5-dichlorophenyl)thio]acetamido]-3-(2-amino-1,3,4-thiadiazol-2-yl)thiomethyl-3-cephem-4-carboxylicacid.
 4. The compound defined in claim 2 wherein the compound is7-[2[(2,5-dichlorophenyl)thio]acetamido]-3-(2-oxazolyl)thiomethyl-3-cephem-4-carboxylicacid.
 5. The compound defined in claim 2 wherein the compound is7-[2[(2,5-dichlorophenyl)thio]acetamido]-3-(5-methyl-1,3,4-oxadiazol-2-yl)thiomethyl-3-cephem-4-carboxylicacid.